Dietary intake of highly polyunsaturated fats is a major source of lipid hydroperoxides in the intestinal lumen. In the preceding grant period, we have established that the intestinal elimination of luminal hydroperoxides is governed by the mucosal glutathione (GSH) antioxidant status. Under conditions of high peroxide intake or compromised GSH status, high concentrations of lipid hydroperoxides can persist in the gut lumen. Since lipid peroxides can initiate degenerative processes via oxyradical formation, constant exposure of the intestinal epithelium to luminal lipid peroxides could impair oxidant detoxication, compromise enterocyte function independent of cell injury and promote development of gut pathologies, including inflammation and cancer. We hypothesize that persistent peroxide challenge at sublethal levels causes a highly oxidized state in the small intestine that compromises mucosal thiol redox balance, impairs GSH-dependent systems and interferes with regulation of intestinal cell death. The overall objectives of the current proposal are two-fold: (a) to understand the cellular and biochemical alterations in intestinal redox/antioxidant status and in intestinal programmed cell death (apoptosis) that are associated with prolonged exposure of the intestinal epithelium to luminal peroxidized lipids, and (b) to determine the role of exogenous GSH supplementation in the preservation of these critical functions. Three specific aims are proposed. AIM 1 will test the hypothesis that persistent exposure of the small intestine to subtoxic levels of peroxidized lipids causes intestinal oxidative stress and inflammation and alters mucosal antioxidant capacity, and this altered metabolic status compromises cellular detoxication of hydroperoxides. AIM 2 will test the hypothesis that exogenous GSH supplementation restores intestinal GSH antioxidant status, decreases tissue oxidative stress and inflammation, and enhances cellular detoxication of hydroperoxides. AIM 3 will test the hypothesis that oxidative stress associated with persistent exposure of the small intestine to subtoxic levels of peroxidized lipids perturbs intestinal apoptosis and GSH supplementation maintains normal cell turnover. The overall strategy is to employ a combination of in vivo (whole animal) and in vitro (isolated cells and purified nuclei) approaches to define the physiological and biochemical processes in lipid hydroperoxide-mediated changes in intestinal redox status, regulation of mucosal thiol and antioxidant balance, and control of intestinal cell death. The results of these findings will have considerable impact on our understanding of dietary oxidants and thiol redox balance on intestinal physiology. The data will also provide useful information regarding the potential use of oral GSH in preserving metabolic integrity of the intestinal epithelium. The long-term goal is to provide a knowledge base to support future clinical efforts to promote tissue antioxidant defenses via dietary or drug therapy.